Abstract
Calcium-dependent cell death occurs in neurodegenerative diseases and ischemic or traumatic brain injury. We analyzed whether thioureylenes can act in a neuroprotective manner by pharmacological suppression of calcium-dependent pathological pathways. In human neuroblastoma (SK-N-SH) cells, thioureylenes (thiopental, carbimazole) inhibited the calcium-dependent neuronal protein phosphatase (PP)-2B, the activation of the proapoptotic transcription factor nuclear factor of activated T-cells, BAD-induced initiation of caspase-3, and poly-(ADP-ribose)-polymerase cleavage. Caspase-3-independent cell death was attenuated by carbimazole and the protein kinase C (PKC) δ inhibitor rottlerin by a PP-2B-independent mechanism. Neuroprotective effects were mediated by the redox-active sulfur of thioureylenes. Furthermore, we observed that the route of calcium mobilization was differentially linked to caspase-dependent or independent cell death and that BAD dephosphorylation did not necessarily induce intrinsic caspase activation. In addition, a new 30- to 35-kDa caspase-3 fragment with an unknown function was identified. In organotypic hippocampal slice cultures, thioureylenes inhibited caspase-3 activation or reduced N-methyl-d-aspartate and kainic acid receptor-mediated cell death that was independent of caspase-3. Because prolonged inhibition of caspase-3 resulted in caspase-independent cellular damage, different types of cell death must be taken under therapeutic consideration. Here we show that thioureylenes in combination with PKCδ inhibitors might represent a promising therapeutic approach to attenuate neuronal damage.
Footnotes
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B.H. and B.H.J.P. contributed equally to this work.
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ABBREVIATIONS: NMDA, N-methyl-d-aspartate; CaM, calmodulin; CsA, cyclosporin A; LDH, lactate dehydrogenase; PARP, poly(ADP-ribose) polymerase; PKC, protein kinase C; PP, protein phosphatase; Z-VAD, benzyloxycarbonyl-Val-Ala-Asp; DEVD, Asp-Glu-Val-Asp; NFAT, nuclear factor of activated T cells; ER, endoplasmic reticulum; DTT, dithiothreitol; TOTEX buffer, HEPES/NaCl/Nonidet P-40/MgCl2/EDTA/EGTA/DTT/phenylmethylsulfonyl fluoride/aprotinin; MK-801, 5H-dibenzo[a,d]cyclohepten-5,10-imine (dizocilpine maleate); Gö6976, 12-(2-cyanoethyl)-6,7,12,13-tetrahydro-13-methyl-5-oxo-5H-indolo(2,3-a)pyrrolo(3,4-c)-carbazole; A23187, calcimycin.
- Received September 23, 2008.
- Accepted December 22, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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